Bactericidal/permeability-increasing protein (BPI) neutralizes bactericidal lipopolysaccharides (LPS) as described in Marta et al. U.S. Pat. No. 5,089,274; Dentener et al. (Oct. 15, 1993) J Immunol 151: 4258-4265; Heumann et al. (1993) J Infectious Diseases 167:1351-1357 and Capadici et al. (January 1994) Infection & Immunity. 62(1):259-265. Bactericidal LPS is a powerful stimulator of the inflammatory response in vivo. In vitro LPS activation of cellular and humoral components of the inflammatory response is blocked by BPI.
Many of the patho-physiologic effects of endotoxin appear to be mediated by the activation of mononuclear cells with the subsequent generation of the proinflammatory cytokines, TNF-alpha and interleukin-1. Macrophage responsiveness to endotoxin varies over time and depends upon several factors (Morrison, D. C. and J. L. Ryan (1987) Ann. Rev. Med. 38:417-432). Endotoxin-signaling pathways within the macrophage are complex and not entirely understood. However, it has become increasingly apparent that endotoxin signaling is initiated with its attachment to the CD14 antigen on the macrophage cell surface (Wright et al. (1990) Science 249:1431-1433). Endogenous proteins which facilitate the delivery of endotoxin to the macrophage cell surface, such as LPS-binding protein (LBP) (Schumann et al (1990) Science 249:1429-1431) stimulate the monocyte/macrophage cell line to produce proinflammatory cytokines and activate a cascade of host-derived inflammatory mediators.
In contrast, BPI blocks endotoxin delivery to the CD14 antigen (Marra et al (1990) J. Immunol. 144:662-666; Marra et al. (1.992) J. Immunol. 148:537). BPI binding to endotoxin has been shown to attenuate cytokine release by mononuclear cells and to inhibit endotoxin mediated activation of neutrophils (Marra et al. (1990), supra; Marra et al. (1992), supra; Weiss et al (1992) J. Clin. Invest. 90:1122-1130; Heumann et al. (1993) J. Infect. Dis. 176:1351-1357).
Most of the various activities of endotoxin have been attributed to the lipid A domian of LPS (Morrison and Ulevitch (1978) Amer. J. Pathol. 93:525). Marra et al. have shown that BPI binds the lipid A domain of endotoxin, thereby preventing subsequent activation of neutrophils (Marra et al. (1990) J. Immunol 144:662). Capodici eta. have reported that BPI blocks the effects of isolated LPS and LPS on Proteus mirabills and E. Coli, which only have the lipid A component in common (Infect. Immun. (1994) 62:25-65).
An example of an in vitro system used to study LPS-mediated cellular activation in vitro is the release of tumor necrosis factor (TNF) and interleukin-1 (IL-1) from macrophages in response to LPS exposure. TNF and IL-1 are cytokines which stimulate fibroblast growth. (LeJ. Vilcek, J. (1987) Lab Invest 56:234). TNF is a potent stimulator of neutrophils.
Polysaccharides other than LPS have also been reported to have immunostimulating activity including stimulation of macrophage function and TNF release. For example, Otterlei et al. (May 1993) Infection and Immunity 61(5):1917-1925, describe the similarity of action of defined polysaccharides and lipopolysaccharides in TNF production.
Entrapment within alginate gel with divalent cation (calcium, barium, strontium or the like) is a widely used technique for immobilization of living cells (Scott, C. D., (1987) Enzyme Microb Technol 9:66). Microencapsulation of hormone-producing cells in calcium alginate has been used for treatment of (1) diabetes mellitus (Lim et al., (1980) Science 21:908; O'Shea et al., (1984) Biochem Biophys Acta 804:133; Goosen et al., (1985) Biotech Bioeng 27:146; Sun et al., "Topics in pharmaceutical science", (1985) Amsterdam, Elsevier Science Publishers B.V. pp.93; Sun, A. M., (1987) Trans Am Soc Artif Organs 33787; Sun et al., (1987) CRC Crit Rev Ther Drug Carrier Sys 4: 1-12; Tze et al., ( 1982) Transplantation 33:563; and Soon-Shiong et al., (1992) Transplantation 54:769-774; (2) liver diseases (Miura et al., (1986) Artif Organs 10:460 and Tompkins et al., (1988) Biotechnol Bioeng 31:11 ); and (3) parathyroid diseases (Darquay et al., (1987) Trans Am Soc Artif Inter Organs 33:365).
Alginates are polysaccharides composed of homopolymeric gel-forming regions of beta-D-mannuronic (M) and alpha-L-glucuronic (G) acid also called "M-blocks" and "G-blocks", respectively, interspaced with regions of mixed sequence ("M-G-blocks") .
The main problem in vivo with alginate capsules is that they induce macrophages and like cells to release cytokines which cause fibroblast overgrowth resulting in failure of encapsulated cells (Sun, A. M., (1987) Trans Am Soc Artif Organs 33:787). Alginates simulate macrophages to release cytokines such as TNF, IL-1, and IL-6. The M-block and MG-blocks but not G-blocks strongly induce cytokine production (Otterlei et al, (1991) J Immunotherapy 10:286-291). If alginates are to be used as a matrix for implanted materials, such as hormone-producing cells, it is important that they do not cause inflammatory reactions.
According to Soon-Shiong et al, "Investigators have long sought to reverse insulin-dependent diabetes by a simple injection of immuno-protected insulin-secreting cells without immunosuppression" (Soon-Shiong et al, (1992) Transplantation, 54:769).
Encapsulation of transplanted insulin-producing (islet) cells provides a method to prevent direct exposure of the host's immune system to the transplanted cells, and this decreases the requirement for immunosuppressants.
Up until the present invention, there has been no suggestion that BPI could have neutralizing activity against molecules other than LPS which can activate inflammatory cells in vitro or in vivo. Data first provided in this application demonstrate that BPI inhibits human monocyte activation (e.g., TNF production) induced by mannuronic acid polymers. Thus, it has been discovered that BPI is useful in preventing and arresting a deleterious inflammatory response to alginate-encapsulated cells for transplant.
In contrast to the prior art, the present invention has demonstrated that BPI can inhibit non-LPS-caused immunostimulation, such as stimulation of macrophage function and TNF release. Thus, BPI has a novel and distinct use in the therapeutic and prophylactic treatment of non-LPS-stimulated inflammation. Furthermore, the invention provides a composition comprising BPI and a non-lipopolysaccharide, such as an alginate gel, which composition inhibits inflammation ordinarily caused by the alginate. Such a composition represents an improvement over the state of the art.